Autism spectrum disorders (ASD), depression and schizophrenia are complex psychiatric conditions involving serotonin (5-HT) dysfunction with a range of symptoms which prominently include impaired social behavior. Drug interventions for these disorders, such as risperidone (Risperdal) and fluoxetine (Prozac), commonly enhance 5-HT neurotransmission or mimic its downstream effects, but these interventions often do not improve social behavior, particularly in patients with gene polymorphisms impairing 5-HT transporter (SERT) function. SERT tightly controls the strength and duration of 5-HT neurotransmission by high affinity uptake of 5-HT from extracellular fluid in brain, but a role for organic cation transporter 3 (OCT3) in 5-HT uptake is emerging as an important mechanism contributing to regulation of extracellular levels of 5-HT in brain. These new findings suggest OCT3 might be a novel target for therapeutic intervention to improve social behavior. Consistent with this idea, we found that acute OCT3 blockade increased social behavior in socially-impaired BTBR mice. The studies we propose here will build on these new findings using SERT knock-out (-/-) mice, which like BTBR mice, are less social than their wild-type (SERT +/+) counterpart. Importantly, SERT-/- mice have increased OCT3 expression and function relative to +/+ mice, making them a unique and useful tool for exploring the effects of OCT3 blockade on 5-HT neurotransmission and social behavior. We hypothesize that the OCT3 blocker, 1,12-diethyl-2,22-cyanine iodide (decynium-22 (D-22)), will improve social behavior in SERT-/- mice more effectively than risperidone, while fluoxetine, which blocks SERT, will have no effect in SERT -/- mice. We expect, however, that D-22 administered in combination with risperidone will produce the greatest increase in social behavior in SERT -/- mice. D-22's effects on SERT +/+ mice, which express both SERT and OCT3 and are highly sociable relative to other strains, will be of interest for use of OCT3 blockade to other ends in broader patient populations. Behavioral data will be correlated with D-22 inhibition of [3H] 5-HT uptake into synaptosomes prepared from hippocampus. Serotonin is a substrate for several biogenic amine transporters, so measuring its uptake in SERT -/- mice, along with selective blockade and control experiments to account for contributions from other transporters expressed in hippocampus, will provide important new information about the relative roles of SERT, OCT3 and other monoamine transporters in 5-HT uptake. Finally, to confirm that D- 22 is exerting its effects centrally, we will measure brain concentration of behaviorally active doses of D-22. Extremely little is known about the role of OCT3 in brain and even less about the effects of systemically administered D-22. Therefore, these fundamental experiments are an essential first step in establishing the potential of OCT3 as a novel target for therapeutic intervention in the treatment of social dysfunction prominent in many psychiatric disorders. PUBLIC HEALTH RELEVANCE: Dysfunction of the serotonin neurotransmitter system has been strongly implicated in the etiology of autism, schizophrenia and depression, and may also underlie impaired social interaction common to these disorders. The proposed studies examine a novel target for pharmaceutical intervention to regulate 5-HT neurotransmission and improve sociability, the organic cation transporter 3 (OCT3), which like the serotonin transporter (SERT, the target of Prozac), removes serotonin from extracellular fluid in the brain to stop neurotransmission. Blockade of OCT3 may prove to be a more uniformly useful mechanism to enhance 5-HT transmission, particularly in patients with SERT gene polymorphisms that make drugs like Prozac less effective.